An assay based on inhibition of microlymphocytoxicity has been developed for monitoring purification of HLA-encoded antigens. Normal methods are not applicable to cell lysates of purified fractions so limited information is available on allospecificities of isolated molecules. The new assay has been successfully used to shed light on the irrelationships of monoclonal antibodies and alloantisera for class II MHC antigens, particularly 33.1. The map order of genes within the HLA complex were analyzed by studying genomic DNA restriction enzyme polymorphisms in seven families that were known to include individuals inheriting recombinant HLA haplotypes. Cloned DNA for class I, class II, (DRAlpha, DRBeta, DCAlpha, DCBeta and SBBeta) and class III (C4) MHC genes were used as probes in Southern blotting analysis. Polymorphism in the size of restriction fragments from the various MHC gene were observed with certain enzymes and could be assigned to haplotypes in the families. The results to date indicate that the genomic restriction enzyme polymorphisms correlate with the assignments made by classical typing techniques.